Fermentative Production of N-Methylglutamate From Glycerol by Recombinant Pseudomonas putida

Mindt, Melanie; Walter, Tatjana; Risse, Joe Max; Wendisch, Volker F.

doi:10.3389/fbioe.2018.00159

Cited by 28 publications

(30 citation statements)

References 69 publications

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“…Many studies deal with the heterologous production by P. putida KT2440. Production of biosurfactants, such as hydroxyalkanoyloxy alkanoates (HAA), mono-rhamnolipids, and di-rhamnolipids (Tiso et al, 2017;Wittgens et al, 2017, terpenoids (zeaxanthin and β-carotene) (Beuttler et al, 2011;Loeschcke et al, 2013), amino acid-derived compounds (e.g., phenazines) (Schmitz et al, 2015b), polyketides/nonribosomal peptides (e.g., flaviolin, prodigiosin) (Gross et al, 2006;Loeschcke et al, 2013;Domröse et al, 2015), and Nmethylglutamate (Mindt et al, 2018) was demonstrated. These examples show how diverse the production spectrum in P. putida KT2440 can be.…”

Section: Discussionmentioning

confidence: 99%

Comparison of Three Xylose Pathways in Pseudomonas putida KT2440 for the Synthesis of Valuable Products

Bator

Wittgens

Rosenau

et al. 2020

Front. Bioeng. Biotechnol.

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Pseudomonas putida KT2440 is a well-established chassis in industrial biotechnology. To increase the substrate spectrum, we implemented three alternative xylose utilization pathways, namely the Isomerase, Weimberg, and Dahms pathways. The synthetic operons contain genes from Escherichia coli and Pseudomonas taiwanensis. For isolating the Dahms pathway in P. putida KT2440 two genes (PP_2836 and PP_4283), encoding an endogenous enzyme of the Weimberg pathway and a regulator for glycolaldehyde degradation, were deleted. Before and after adaptive laboratory evolution, these strains were characterized in terms of growth and synthesis of mono-rhamnolipids and pyocyanin. The engineered strain using the Weimberg pathway reached the highest maximal growth rate of 0.30 h −1. After adaptive laboratory evolution the lag phase was reduced significantly. The highest titers of 720 mg L −1 mono-rhamnolipids and 30 mg L −1 pyocyanin were reached by the evolved strain using the Weimberg or an engineered strain using the Isomerase pathway, respectively. The different stoichiometries of the three xylose utilization pathways may allow engineering of tailored chassis for valuable bioproduct synthesis.

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Section: Discussionmentioning

confidence: 99%

Comparison of Three Xylose Pathways in Pseudomonas putida KT2440 for the Synthesis of Valuable Products

Bator

Wittgens

Rosenau

et al. 2020

Front. Bioeng. Biotechnol.

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“…These processes differ regarding the substrate spectra of the involved enzymes. For example, anthranilate N-methylation described here as well as N-methylglutamate production established in Pseudomonas putida using N-methylglutamate synthase and γ-glutamylmethylamide synthetase of the methylamine assimilation pathway of Methylobacterium extorquens [13] have narrow substrate spectra (e.g., GMAS from Methylovorus mays also forms γ-glutamylethylamide, also known as theanine [69]) compared with N-alkylation using the imine reductase DpkA of Pseudomonas putida [12]. Several methylated or ethylated amino acids could be produced by C. glutamicum using the wild-type or a mutant version of DpkA and either MMA or ethylamine as substrates [14,34,35].…”

Section: Discussionmentioning

confidence: 99%

“…Fermentation processes using simple mineral salts media have been developed for three different routes for de novo production of N -alkylated amino acids. Two metabolic engineering strategies for reductive alkylamination of 2-oxo acids with monomethylamine that either make use of a C1-assimilation pathway present in methylotrophic bacteria [ 13 ] or of the imine reductase DpkA [ 14 ] have been established. S -Adenosyl- l -methionine (SAM)-dependent methylation of aromatic amino acids by N -methyltransferases has also been described [ 15 ].…”

Section: Introductionmentioning

confidence: 99%

Fermentative N-Methylanthranilate Production by Engineered Corynebacterium glutamicum

et al. 2020

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The N-functionalized amino acid N-methylanthranilate is an important precursor for bioactive compounds such as anticancer acridone alkaloids, the antinociceptive alkaloid O-isopropyl N-methylanthranilate, the flavor compound O-methyl-N-methylanthranilate, and as a building block for peptide-based drugs. Current chemical and biocatalytic synthetic routes to N-alkylated amino acids are often unprofitable and restricted to low yields or high costs through cofactor regeneration systems. Amino acid fermentation processes using the Gram-positive bacterium Corynebacterium glutamicum are operated industrially at the million tons per annum scale. Fermentative processes using C. glutamicum for N-alkylated amino acids based on an imine reductase have been developed, while N-alkylation of the aromatic amino acid anthranilate with S-adenosyl methionine as methyl-donor has not been described for this bacterium. After metabolic engineering for enhanced supply of anthranilate by channeling carbon flux into the shikimate pathway, preventing by-product formation and enhancing sugar uptake, heterologous expression of the gene anmt encoding anthranilate N-methyltransferase from Ruta graveolens resulted in production of N-methylanthranilate (NMA), which accumulated in the culture medium. Increased SAM regeneration by coexpression of the homologous adenosylhomocysteinase gene sahH improved N-methylanthranilate production. In a test bioreactor culture, the metabolically engineered C. glutamicum C1* strain produced NMA to a final titer of 0.5 g·L−1 with a volumetric productivity of 0.01 g·L−1·h−1 and a yield of 4.8 mg·g−1 glucose.

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“…Notably, sustainable fermentative production of N -methylated amino acids was established in 2018 using two independent pathways. Pathway interception of the C1 assimilation pathway of Methylobacterium extorquens and implementation of the first two genes coding for γ-glutamylmethylamide synthetase and N -methylglutamate synthase enabled glycerol-based production of N -methylglutamate in Pseudomonas putida KT2440 (Mindt et al, 2018b). The second pathway for production of N -methylated amino acids exploited the side activity of the imine reductase DpkA from Pseudomonas putida .…”

Section: Introductionmentioning

confidence: 99%

Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida

Mindt

Hannibal

Heuser

et al. 2019

Front. Bioeng. Biotechnol.

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Sarcosine, an N-methylated amino acid, shows potential as antipsychotic, and serves as building block for peptide-based drugs, and acts as detergent when acetylated. N-methylated amino acids are mainly produced chemically or by biocatalysis, with either low yields or high costs for co-factor regeneration. Corynebacterium glutamicum, which is used for the industrial production of amino acids for decades, has recently been engineered for production of N-methyl-L-alanine and sarcosine. Heterologous expression of dpkA in a C. glutamicum strain engineered for glyoxylate overproduction enabled fermentative production of sarcosine from sugars and monomethylamine. Here, mutation of an amino acyl residue in the substrate binding site of DpkA (DpkAF117L) led to an increased specific activity for reductive alkylamination of glyoxylate using monomethylamine and monoethylamine as substrates. Introduction of DpkAF117L into the production strain accelerated the production of sarcosine and a volumetric productivity of 0.16 g L−1 h−1 could be attained. Using monoethylamine as substrate, we demonstrated N-ethylglycine production with a volumetric productivity of 0.11 g L−1 h−1, which to the best of our knowledge is the first report of its fermentative production. Subsequently, the feasibility of using rice straw hydrolysate as alternative carbon source was tested and production of N-ethylglycine to a titer of 1.6 g L−1 after 60 h of fed-batch bioreactor cultivation could be attained.

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Fermentative Production of N-Methylglutamate From Glycerol by Recombinant Pseudomonas putida

Cited by 28 publications

References 69 publications

Comparison of Three Xylose Pathways in Pseudomonas putida KT2440 for the Synthesis of Valuable Products

Comparison of Three Xylose Pathways in Pseudomonas putida KT2440 for the Synthesis of Valuable Products

Fermentative N-Methylanthranilate Production by Engineered Corynebacterium glutamicum

Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida

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